Recent studies have shown that there exists a family of mammalian protein kinases structurally and functionally related to the yeast cell cycle regulatory kinase cdc-2. Two members of the cdc2 family, cdk1 and cdk2, have been identified in mammalian cells. Cdk1 kinase regulates the progression from G2 to M, and cdk2 has been proposed to regulate the progression from G1 to S phase. We have cloned and structurally characterized an additional member of the cdc2 kinase family, cdk 5, with 58% amino acid sequence identity to mouse cdk 1 and 60% identity to human cdk2. In contrast to either cdk 1 or cdk2, cdk 5 is expressed at high levels in terminally differentiated neurons. Moreover, many cultured cell lines express high levels of cdk 5 mRNA as well as cdk 1 and cdk2 kinase. We are exploring the role of cdk 5 in cell cycle regulation of mitotically active cells and human tumors. We have generated an antipeptide antiserum specific for cdk 5, which can specifically immunoprecipitate cdk 5 kinase from mammalian cell extracts, and binds specifically to cdk 5 in Western blotting studies. Using this antiserum, we have established that cultured human tumor cells express enzymatically active cdk 5 kinase. In addition, cdk 5 and cyclin D, a regulatory factor needed for kinase activity, have been co-expressed in E. coli. Characterization of the enzymatic properties of the kinase produced in bacteria is currently underway.